5-HT1D ( pKi = 7.9 ); 5-HT1A ( pKi = 7.7 ); 5-HT2B ( pKi = 7.4 ); 5-HT2A ( pKi = 6.6 ); 5-HT7 ( pKi = 6.3 )

体外活性：BRL-15572 对 h5-HT1D 受体表现出高亲和力和选择性。 BRL-15572 对 h5-HT1D 的亲和力比 5-HT1B 受体高 60 倍。 BRL-15572 与 h5-HT1B 和 h5-HT1D 受体结合，pKB 分别小于 6 和 7.1。 BRL-15572 刺激两种细胞系中的 [35S]GTP γ S 结合，其效力与其在 h5-HT1B 和 h5-HT1D 受体表达细胞系中的受体结合亲和力相关。 BRL-15572 揭示了对 5-HT1A、5-HT1B、5-HT1E、5-HT1F、5-HT2A、5-HT2B、5-HT2C、5-HT6 和 5-HT7 的受体结合亲和力，pKi 为 7.7、6.1、分别为 5.2、6.0、6.6、7.4、6.2、5.9 和 6.3。在 h5-HT1D 细胞系中，两种 BRL-15572 (1 µM) 均会改变 5-HT 浓度响应曲线，pKB 分别为 7.1。 BRL-15572 对人类 5-HT1A 和 5-HT2B 受体具有中等高的亲和力。在人心耳中，电诱发的氚溢出被 5-HT 抑制，其方式容易受到 BRL-15572（300 nM；h5-HT1D 受体 Ki 的 23 倍）的拮抗作用。 5-HT 对 K+ 引起的谷氨酸溢出的抑制作用被 h5-HT1D 受体配体 BRL-15572 拮抗。 BRL-15572 (1 μM) 无法改变 5-HT 对自身受体调节 [3H]5-HT 释放的影响。选择性 5-HT1D/1B 受体拮抗剂 BRL 15572 抑制激动剂 L-694 247 的作用。细胞测定：[35S]GTPγS 结合研究。在表达 h5-HT1B 或 h5-HT1D 受体的 CHO 细胞中进行 [35S]GTPγS 结合研究。简而言之，将 1 × 106 个细胞的膜在含有 GDP 的 HEPES 缓冲液（HEPES [20 mM]、MgCl2 [3 mM]、NaCl [100 mM]、抗坏血酸 [0.2 mM]）中预孵育 30 分钟。 (10 µ M)，有或没有 BRL-15572。通过添加 10 µL [35S]GTPγS（100 pM，测定浓度）开始反应，然后在 30°C 下进一步孵育 30 分钟。在添加细胞之前，通过添加未标记的 GTPγS (10 µM) 来确定非特异性结合。使用 Whatman GF/B 级过滤器快速过滤，然后用冰冷的 HEPES 缓冲液洗涤五次，从而终止反应。放射性通过液体闪烁光谱法测定。

在糖尿病大鼠中，给予选择性 5-HT1D 受体拮抗剂 BRL-15572 (2 mg/kg) 不会改变迷走神经电刺激引起的 HR 降低。 L-694,247 (50 μg/kg)（一种非啮齿动物 5-HT1B 和 5-HT1D 受体的选择性激动剂）在用 BRL-15572 预处理后对迷走神经诱导的心动过缓的影响并不明显。

尽管h5-HT1B和h5-HT1D受体氨基酸序列之间只有适度的同源性，但这些受体显示出非常相似的药理学。迄今为止，很少有化合物能区分这些受体亚型和那些具有一定选择性的化合物，如酮色林，对其他5-HT受体亚型具有更大的亲和力。我们现在报告了两种化合物，SB-216641（N-[3-（2-二甲氨基）乙氧基-4-甲氧基苯基]-2'-甲基-4'-（5-甲基-1,2,4-恶二唑-3-基）-（1,1'-联苯）-4-甲酰胺）和BRL-155723-[4-（3-氯苯基）哌嗪-1-基]-1,1-二苯基-2-丙醇），它们分别对h5-HT1B和h5-HT1D受体显示出高亲和力和选择性。在CHO细胞中表达的人受体的受体结合研究中，SB-216641对h5-HT1B受体具有高亲和力（pKi=9.0），对h5-HT1D受体的亲和力低25倍。相比之下，BRL-15572对h5-HT1D（pKi=7.9）的亲和力比5-HT1B受体高60倍。在豚鼠纹状体的天然组织5-HT1B受体上测定了这些化合物的类似亲和力。在[35S]GTPγS结合试验和重组h5-HT1B和h5-HT1D受体上的cAMP积累试验中测量了SB-216641和BRL-15572的功能活性。这两种化合物在这些高受体表达系统中都是部分激动剂，其效力和选择性与其受体结合亲和力相关。在cAMP积累测定中，化合物的pK（B）测量结果再次与受体结合亲和力相关（SB-216641，pK（B）=9.3和7.3BRL-15572，pK（B）=<6，h5-HT1B和h5-HT1D受体分别为7.1）。这些化合物将成为表征5-HT1B和5-HT1D受体介导反应的有用药理学试剂[1]。

[35S]GTPγS 结合研究。 [³⁵S]GTPγS 结合研究是在表达 h5-HT_1B 或 h5-HT_1D 受体的 CHO 细胞中进行的。总之，1 × 10⁶ 细胞膜在 HEPES 缓冲液中预孵育（HEPES [20 mM]、MgCl₂ [3 mM]、NaCl [100 mM]、抗坏血酸 [ 0.2 mM]），含 GDP (10 µ M)，含或不含 BRL-15572，30°C 30 分钟。以 10 µL 增量添加 100 pM 测定浓度的 [³⁵S]GTPγS 以启动反应，然后在 30°C 下再孵育 30 分钟。非特异性结合的测定是通过首先添加未标记的 GTPγS (10 µM)，然后添加细胞来实现的。使用 Whatman GF/B 级过滤器快速过滤掉反应物，然后进行五次冰冷 HEPES 缓冲液洗涤。液体闪烁光谱用于测量放射性。

Human cerebral cortical slices and synaptosomes, guinea-pig cerebral cortical slices and human right atrial appendages were used to study the effects of SB-216641, a preferential h5-HT1B receptor ligand, and of BRL-15572, a preferential h5-HT1D receptor ligand, on the presynaptic h5-HT1B and h5-HT1B-like autoreceptors in the human and guinea-pig brain preparations, respectively, and on the presynaptic h5-HT1D heteroreceptors in the human atrium. The brain preparations, preincubated with [3H]serotonin ([3H]5-HT), and the segments of atrial appendages, preincubated with [3H]noradrenaline, were superfused with modified Krebs' solution and tritium overflow was evoked electrically (human and guinea-pig cerebral cortex slices and human atrial appendages) or by high K+ (human cerebral cortex synaptosomes). The electrically evoked tritium overflow from guinea-pig cerebral cortex slices was reduced by the 5-HT receptor agonist 5-carboxamidotryptamine (5-CT). This effect was not modified by BRL-15572 (2 microM; concentration 154 times higher than its Ki at h5-HT1D receptors) but was antagonized by SB-216641 (0.1 microM; concentration 100 times higher than its Ki at h5-HT1B receptors; apparent pA2 8.45). SB-216641 (0.1 microM) by itself facilitated, whereas BRL-15572 (2 microM) did not affect, the evoked overflow. In human cerebral cortex slices SB-216641 (0.1 microM) also facilitated, and BRL-15572 (2 microM) again failed to affect, the electrically evoked tritium overflow. In human cerebral cortical synaptosomes, 5-CT reduced the K+-evoked tritium overflow. This response was unaffected by BRL-15572 (300 nM) but antagonized by SB-216641 (15 nM; drug concentrations 23 and 15 times higher than their Ki at h5-HT1D and h5-HT1B receptors, respectively). Both drugs, given alone, did not modify the K+-evoked tritium overflow. In human atrial appendages, the electrically evoked tritium overflow was inhibited by 5-HT in a manner susceptible to antagonism by BRL-15572 (300 nM; 23 times Ki at h5-HT1D receptors) but not by SB-216641 (30 nM; 30 times Ki at h5-HT1B receptors). Both drugs by themselves did not change the electrically evoked tritium overflow. In conclusion, SB-216641 behaves as a preferential antagonist at native human 5-HT1B receptors and BRL-15572 as a preferential antagonist at native human 5-HT1D receptors. These compounds are clearly useful tools for the differentiation between human 5-HT1B and 5-HT1D receptors in functional studies.[2]

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Dissolved in 20% propylene glycol; 1 mg/kg, 2 mg/kg; i.v. injection

Male Wistar rats with diabetes

[1]. Naunyn Schmiedebergs Arch Pharmacol. 1997 Sep;356(3):312-20.

[2]. Naunyn Schmiedebergs Arch Pharmacol. 1997 Sep;356(3):321-7.

[3]. Br J Pharmacol. 1999 Feb;126(3):607-12.

[4]. Clin Exp Pharmacol Physiol. 2005 Oct;32(10):894-900.

[5]. Clin Exp Pharmacol Physiol. 2007 Nov;34(11):1199-206.

[6]. Naunyn Schmiedebergs Arch Pharmacol. 2004 Jul;370(1):46-53.

1. In the present study, we investigated how alloxan-induced diabetes affects the ability of 5-hydroxytryptamine (5-HT) to modulate bradycardia induced in vivo by electrical stimulation of the vagus nerve in pithed rats. We also analysed the type and/or subtype of 5-HT receptors involved. 2. Diabetes was induced in male Wistar rats with a single injection of alloxan (150 mg/kg, s.c.). Four weeks later, rats were anaesthetized, pretreated with atenolol and pithed. Electrical stimulation (3, 6 and 9 Hz) of the vagus nerve resulted in frequency dependent decreases in heart rate (HR). 3. In diabetic rats, intravenous bolus administration of high doses of 5-HT (100 and 200 microg/kg) increased the bradycardia induced by vagal electrical stimulation. Similarly, low doses (10 microg/kg) of the 5-HT(1/7) receptor agonist 5-carboxamidotryptamine (5-CT), increased vagally induced bradycardia. However, at high doses (50, 100 and 150 microg/kg), 5-CT reduced the bradycardia. Attenuation of the vagally induced bradycardia evoked by the higher doses of 5-CT was reproduced by L-694,247 (50 microg/kg), a selective agonist for the non-rodent 5-HT(1B) and 5-HT(1D) receptors. Enhancement of the vagally induced bradycardia elicited by low doses of 5-CT was reproduced by the selective 5-HT(1A) receptor agonist 8-hydroxydipropylaminotretalin hydrobromide (8-OH-DPAT; 50 microg/kg). These stimulatory and inhibitory actions on vagal stimulation-induced bradycardia in diabetic rats were also observed after administration of exogenous acetylcholine. 4. Vagally induced bradycardia in diabetic rats was not affected by administration of the selective 5-HT(2) receptor agonist alpha-methyl-5-HT (150 microg/kg), the selective 5-HT(3) receptor agonist 1-phenylbiguanide (150 microg/kg) or the selective 5-HT(1B) receptor agonist CGS-12066B (50 microg/kg). 5. Enhancement of the electrical stimulation-induced bradycardia in diabetic rats caused by 5-CT (10 microg/kg) or 8-OH-DPAT (50 microg/kg) was abolished by the selective 5-HT(2/7) receptor antagonist mesulergine (1 mg/kg) and the selective 5-HT(1A) receptor antagonist WAY-100,635 (100 microg/kg), respectively. Similarly, pretreatment with the non-selective 5-HT(1) receptor antagonist methiothepin (0.1 mg/kg) blocked the inhibitory effect of 5-CT (50 microg/kg) on the bradycardia induced by vagal electrical stimulation in diabetic rats. BRL-15572 (2 microg/kg), a selective 5-HT(1D) receptor antagonist, inhibited the action of L-694,247 (50 microg/kg), a selective agonist for the non-rodent 5-HT(1B) and 5-HT(1D) receptors, on the vagally induced bradycardia. 6. In conclusion, in the present study, experimental diabetes evoked changes in both the nature and 5-HT receptor types/subtypes involved in vagally induced bradycardia.[5]

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It has previously been suggested that ergotamine produces external carotid vasoconstriction in vagosympathectomised dogs via 5-HT1B/1D receptors and alpha2-adrenoceptors. The present study has reanalysed this suggestion by using more selective antagonists alone and in combination. Fifty-two anaesthetised dogs were prepared for ultrasonic measurements of external carotid blood flow. The animals were divided into thirteen groups (n=4 each) receiving an i.v. bolus injection of, either physiological saline (0.3 ml/kg; control), or the antagonists SB224289 (300 microg/kg; 5-HT1B), BRL15572 (300 microg/kg; 5-HT1D), rauwolscine (300 microg/kg; alpha2), SB224289 + BRL15572 (300 microg/kg each), SB224289 + rauwolscine (300 microg/kg each), BRL15572 + rauwolscine (300 microg/kg each), rauwolscine (300 microg/kg) + prazosin (100 microg/kg; alpha1), SB224289 (300 microg/kg) + prazosin (100 microg/kg), SB224289 (300 microg/kg) + rauwolscine (300 microg/kg) + prazosin (100 microg/kg), SB224289 (300 microg/kg) + prazosin (100 microg/kg) + BRL44408 (1,000 microg/kg; alpha2A), SB224289 (300 microg/kg) + prazosin (100 microg/kg)+ imiloxan (1,000 microg/kg; alpha2B), or SB224289 (300 microg/kg) + prazosin (100 microg/kg) + MK912 (300 microg/kg; alpha2C). Each group received consecutive 1-min intracarotid infusions of ergotamine (0.56, 1, 1.8, 3.1, 5.6, 10 and 18 microg/min), following a cumulative schedule. In saline-pretreated animals, ergotamine induced dose-dependent decreases in external carotid blood flow without affecting arterial blood pressure or heart rate. These control responses were: unaffected by SB224289, BRL15572, rauwolscine or the combinations of SB224289 + BRL15572, BRL15572 + rauwolscine, rauwolscine + prazosin, SB224289 + prazosin, or SB224289 + prazosin + imiloxan; slightly blocked by SB224289 + rauwolscine; and markedly blocked by SB224289 + rauwolscine + prazosin, SB224289 + prazosin + BRL44408 or SB224289 + prazosin + MK912. Thus, the cranio-selective vasoconstriction elicited by ergotamine in dogs is predominantly mediated by 5-HT1B receptors as well as alpha2A/2C-adrenoceptor subtypes and, to a lesser extent, by alpha1-adrenoceptors.[6]

C25H29CL3N2O

479.87

478.13

1173022-77-9

BRL-15572 hydrochloride; 1173022-77-9

9891303

White to off-white solid powder

580.7ºC at 760 mmHg

305ºC

2.51E-14mmHg at 25°C

5.459

26.71

3

3

6

31

451

0

0

WPEXRXMQMPOHIO-UHFFFAOYSA-N

InChI=1S/C25H27ClN2O.2ClH/c26-22-12-7-13-23(18-22)28-16-14-27(15-17-28)19-24(29)25(20-8-3-1-4-9-20)21-10-5-2-6-11-21;;/h1-13,18,24-25,29H,14-17,19H2;2*1H

3-[4-(3-chlorophenyl)piperazin-1-yl]-1,1-diphenylpropan-2-ol;dihydrochloride

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

注意： 请将本产品存放在密封且受保护的环境中，避免吸湿/受潮。

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

配方 1 中的溶解度: ≥ 2.08 mg/mL (4.33 mM) (饱和度未知) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将100 μL 20.8 mg/mL澄清DMSO储备液加入400 μL PEG300中，混匀；然后向上述溶液中加入50 μL Tween-80，混匀；加入450 μL生理盐水定容至1 mL。
*生理盐水的制备：将 0.9 g 氯化钠溶解在 100 mL ddH₂O中，得到澄清溶液。

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配方 2 中的溶解度: ≥ 2.08 mg/mL (4.33 mM) (饱和度未知) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将 100 μL 20.8 mg/mL澄清DMSO储备液加入900 μL 20% SBE-β-CD生理盐水溶液中，混匀。
*20% SBE-β-CD 生理盐水溶液的制备（4°C，1 周）：将 2 g SBE-β-CD 溶解于 10 mL 生理盐水中，得到澄清溶液。

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配方 3 中的溶解度: ≥ 2.08 mg/mL (4.33 mM) (饱和度未知) in 10% DMSO + 90% Corn Oil (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将 100 μL 20.8 mg/mL 澄清 DMSO 储备液添加到 900 μL 玉米油中并混合均匀。

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配方 4 中的溶解度: 30% Propylene glycol , 5% Tween 80 , 65% D5W: 20 mg/mL

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请根据您的实验动物和给药方式选择适当的溶解配方/方案：
1、请先配制澄清的储备液(如：用DMSO配置50 或 100 mg/mL母液(储备液));
2、取适量母液，按从左到右的顺序依次添加助溶剂，澄清后再加入下一助溶剂。以 下列配方为例说明 (注意此配方只用于说明，并不一定代表此产品 的实际溶解配方):
10% DMSO → 40% PEG300 → 5% Tween-80 → 45% ddH2O (或 saline);
假设最终工作液的体积为 1 mL, 浓度为5 mg/mL: 取 100 μL 50 mg/mL 的澄清 DMSO 储备液加到 400 μL PEG300 中，混合均匀/澄清；向上述体系中加入50 μL Tween-80，混合均匀/澄清；然后继续加入450 μL ddH2O (或 saline)定容至 1 mL；
3、溶剂前显示的百分比是指该溶剂在最终溶液/工作液中的体积所占比例;
4、 如产品在配制过程中出现沉淀/析出，可通过加热(≤50℃)或超声的方式助溶;
5、为保证最佳实验结果，工作液请现配现用！
6、如不确定怎么将母液配置成体内动物实验的工作液，请查看说明书或联系我们;
7、 以上所有助溶剂都可在 Invivochem.cn网站购买。